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HelpTest ID F822P Hemophilia A F8 Gene, Intron 22 Inversion Mutation Analysis, Prenatal
Useful For
Prenatal testing for hemophilia A when a F8 intron 22 inversion has been identified in a family member
Disease States
- Hemophilia A
Reflex Tests
| Test ID | Reporting Name | Available Separately | Always Performed |
|---|---|---|---|
| CULAF | Amniotic Fluid Culture/Genetic Test | Yes | No |
| CULFB | Fibroblast Culture for Genetic Test | Yes | No |
| MATCC | Maternal Cell Contamination, B | Yes | No |
Testing Algorithm
If amniotic fluid is received, amniotic fluid culture/genetic test will be added at an additional charge.
If chorionic villus specimen is received, fibroblast culture for genetic test will be added at an additional charge.
For any prenatal specimen that is received, maternal cell contamination studies will be added. A maternal whole blood specimen is required to perform this test.
The following algorithms are available:
Reporting Name
HA F8 Int22 Inversion KM, AF or CVSSpecimen Type
VariesAdditional Testing Requirements
Due to its complexity, consultation with the laboratory is required for all prenatal testing; call 800-533-1710 to speak to a genetic counselor.
All prenatal specimens must be accompanied by a maternal blood specimen. Order MATCC / Maternal Cell Contamination, Molecular Analysis, Varies on the maternal specimen.
Shipping Instructions
Advise Express Mail or equivalent if not on courier service
Necessary Information
Hemophilia A Patient Information (T712) is required. Testing may proceed without the patient information, however, the information aids in providing a more thorough interpretation. Ordering providers are strongly encouraged to fill out the form and send with the specimen.
Specimen Required
Results will be reported and telephoned or faxed if requested.
Submit only 1 of the following specimens:
Specimen Type: Amniotic fluid
Container/Tube: Amniotic fluid container
Specimen Volume: 5 to 10 mL
Collection Instructions:
1. Optimal timing for specimen collection is during 14 to 18 weeks of gestation, but specimens collected at other weeks of gestation are also accepted.
2. Discard the first 2 mL of amniotic fluid. If the culture will be performed in conjunction with chromosome analysis and alpha-fetoprotein, a total of approximately 25 to 30 mL will be needed for the combined studies.
Specimen Stability Information: Ambient (preferred) <24 hours/Refrigerated
Additional Information:
1. Place the tubes in a Styrofoam container.
2. Fill remaining space with packing material.
3. Unavoidably, about 1% to 2% of mailed-in specimens are not viable.
4. Bloody specimens are undesirable.
5. If the specimen does not grow in culture, you will be notified within 7 days of receipt.
6. A separate culture charge will be assessed under CULAF / Culture for Genetic Testing, Amniotic Fluid.
7. All prenatal specimens must be accompanied by a maternal blood specimen; order MATCC / Maternal Cell Contamination, Molecular Analysis, Varies on the maternal specimen.
Specimen Type: Chorionic villi
Supplies: CVS Media (RPMI) and Small Dish (T095)
Container/Tube: 15-mL tube containing 15 mL of transport media
Specimen Volume: 20 to 30 mg
Collection Instructions:
1. Collect specimen by the transabdominal or transcervical method.
2. Transfer the chorionic villi specimen to a Petri dish containing transport medium.
3. Using a stereomicroscope and sterile forceps, assess the quality and quantity of the villi and remove any blood clots and maternal decidua.
Specimen Stability Information: Refrigerated (preferred) <24 hours/Ambient
Additional Information:
1. A separate culture charge will be assessed under CULFB / Fibroblast Culture for Biochemical or Molecular Testing. An additional 3 to 4 weeks is required to culture fibroblasts before genetic testing can occur.
2. All prenatal specimens must be accompanied by a maternal blood specimen; order MATCC / Maternal Cell Contamination, Molecular Analysis, Varies on the maternal specimen.
Specimen Type: Confluent cultured cells
Container/Tube: T-25 flask
Specimen Volume: 2 flasks approximately 90% confluent
Collection Instructions: Submit confluent cultured cells from another laboratory.
Specimen Stability Information: Ambient (preferred) <24 hours/Refrigerated
Additional Information: All prenatal specimens must be accompanied by a maternal blood specimen; order MATCC / Maternal Cell Contamination, Molecular Analysis, Varies on the maternal specimen.
Specimen Minimum Volume
Amniotic fluid: See Specimen Required
Chorionic villi: 5 mg
Specimen Stability Information
| Specimen Type | Temperature | Time | Special Container |
|---|---|---|---|
| Varies | Varies | ||
Reject Due To
All specimens will be evaluated at Mayo Clinic Laboratories for test suitability.Genetics Test Information
This test detects the intron 22 inversion within the F8 gene. The intron 22 inversion variant accounts for approximately 45% of variants associated with severe hemophilia A.
Intron 22 inversion known variant analysis on a prenatal specimen can only be performed when there is a known intron 22 inversion in the family.
Clinical Information
Hemophilia A (HA) is caused by a deficiency of clotting factor VIII (FVIII). HA is an X-linked recessive bleeding disorder that affects approximately 1 in 5000 male individuals. Male patients are typically affected with bleeding symptoms, whereas female carriers generally do not have bleeding symptoms but are at risk of having affected sons. Rarely, approximately 10% of female carriers have FVIII activity levels below 35% and are at risk for bleeding.
Bleeding, the most common clinical symptom in individuals with HA, correlates with FVIII activity levels. FVIII activity levels below 1% are associated with severe disease, 1% to 5% activity with moderate disease, and 5% to 40% with mild disease. In male patients with severe deficiency, spontaneous bleeding may occur. In individuals with mild HA, bleeding may occur only after surgery or trauma.
FVIII is encoded by the factor VIII (F8) gene. Approximately 98% of patients with a diagnosis of HA are found to have a variant in F8 (ie, intron 1 and 22 inversions, point mutations, insertions, and deletions). The intron 1 inversion variant accounts for approximately 5% of variants associated with severe HA. These inversions are typically not identified in patients with mild or moderate HA.
Intron 22 inversion known variant analysis on a prenatal specimen can only be performed when there is a known intron 22 inversion in the family.
It is recommended that the F8 variant be confirmed in the affected male patient or obligate female carrier prior to testing at-risk individuals. Affected male patients are identified by FVIII activity (F8A / Coagulation Factor VIII Activity Assay, Plasma) and clinical evaluation, while obligate female carriers are identified by family history assessment. If the intron inversion assays do not detect an inversion in these individuals, additional analysis (ie, F8 sequencing) may be able to identify the familial variant. Of note, not all women with an affected son are germline carriers of a F8 variant, as de novo variants in F8 do occur. Approximately 20% of mothers of isolated cases do not have an identifiable germline F8 variant. Importantly, there is a small risk for recurrence even when the familial F8 variant is not identified in the mother of the affected patient due to the possibility of germline mosaicism.
Reference Values
An interpretive report will be provided.
Interpretation
The interpretive report will include assay information, background information, and conclusions based on the test results.
Cautions
Obtaining a medical genetics or hematology (coagulation) consultation prior to ordering is advisable. Molecular genetic or hemophilia center consultation is available for all possible hemophilia A cases and is particularly indicated in complex cases or in situations in which the diagnosis is atypical or uncertain.
This assay detects only F8 intron 22 inversion variants. Thus, a negative result does not exclude the presence of other variants in F8.
The intron 22 inversion variant targeted by this assay is found in approximately 45% of individuals with severe hemophilia A; if an intron 22 inversion has not been already identified in the family, the assay may be uninformative.
Test results should be interpreted in the context of clinical findings, family history, and other laboratory data. Errors in the interpretation of results may occur if the information provided is inaccurate or incomplete.
Clinical Reference
1. Antonarakis SE, Rossiter JP, Young M, et al: Factor VIII gene inversions in severe hemophilia A: results of an international consortium study. Blood. 1995 Sep 15;86(6):2206-2212
2. Rossiter JP, Young M, Kimberland ML, et al: Factor VIII gene inversions causing severe hemophilia A originate almost exclusively in male germ cells. Hum Mol Genet. 1994 Jul;3(7):1035-1039
3. Castaldo G, D'Argenio V, Nardiello P, et al: Haemophilia A: molecular insights. Clin Chem Lab Med. 2007;45(4):450-461
4. Oldenburg J, Rost S, El-Maarri O, et al: De novo factor VIII gene intron 22 inversion in a female carrier presents as a somatic mosaicism. Blood. 2000 Oct 15;96(8):2905-2906
5. Pruthi RK: Hemophilia: a practical approach to genetic testing. Mayo Clin Proc. 2005 Nov;80(11):1485-1499
6. Johnsen JM, Fletcher SN, Huston H, et al: Novel approach to genetic analysis and results in 3000 hemophilia patients enrolled in the My Life, Our Future initiative. Blood Adv. 2017 May 18;1(13):824-834. doi: 10.1182/bloodadvances.2016002923
Method Description
Genomic DNA from amniotic fluid or chorionic villi is digested with restriction enzyme, ligated with T4 DNA ligase, and amplified by polymerase chain reaction with primers specific for the F8 intron 22 inversion variants.(Rosetti LC, Radic CP, Larripa IB, De Brasi CD: Developing a new generation of tests for genotyping hemophilia-causative rearrangements involving int22h and int1h hotspots in the factor VIII gene. J Thromb Haemost. 2008;6:830-836)
Day(s) Performed
Monday through Friday
Report Available
28 to 35 daysTest Classification
This test was developed and its performance characteristics determined by Mayo Clinic in a manner consistent with CLIA requirements. It has not been cleared or approved by the US Food and Drug Administration.CPT Code Information
81403
NY State Approved
YesMethod Name
Inverse Shifting-Polymerase Chain Reaction (IS-PCR)
Forms
1. Hemophilia A Patient Information (T712) is required.
2. New York Clients-Informed consent is required. Document on the request form or electronic order that a copy is on file. The following documents are available:
-Informed Consent for Genetic Testing (T576)
-Informed Consent for Genetic Testing-Spanish (T826)
3. If not ordering electronically, complete, print, and send a Coagulation Test Request (T753) with the specimen.